摘要
目的:研究益肾化浊方(YHP)早期干预对快速衰老模型小鼠学习记忆的影响及其作用机制。方法:将48只3月龄雄性SAMP8小鼠随机分为模型组、YHP低、高剂量组、多奈哌齐组;同月龄SAMR1小鼠24只,随机分为空白组、空白给药组,每组12只。YHP组给予6.24、12.48 g·kg^(-1)YHP,多奈哌齐组给予0.65 mg·kg^(-1)盐酸多奈哌齐,空白组和模型组予生理盐水,每天灌胃1次,连续4周。Morris水迷宫实验评价小鼠空间学习记忆能力。免疫荧光法评估大脑海马区神经元密度及小胶质细胞(MG)极化标记物M1型-诱导型一氧化氮合酶(i NOS)和M2型-精氨酸酶-1(Arg-1)表达情况。酶联免疫吸附测定法(ELISA)检测血清促炎因子白细胞介素-1β(IL-1β)和抑炎因子转化生长因子-β1(TGF-β1)含量。蛋白免疫印迹法(Western blot)检测海马组织β淀粉样蛋白1-42(Aβ1-42)和髓样细胞表达触发受体2(TREM2)/核转录因子-κB(NF-κB)信号通路相关蛋白髓样细胞表达触发受体2(TREM2)、磷酸化(p)-NF-κB p65、磷酸化NF-κB抑制因子激酶β(p-IKKβ)表达情况。结果:与空白组比较,模型组逃避潜伏期显著延长,海马神经元特异性核蛋白(Neu N)表达显著降低,MG的i NOS表达显著增多,Arg-1表达显著降低(P<0.01);血清IL-1β含量显著增多,TGF-β1含量明显降低;海马TREM2表达显著降低,p-NF-κB p65、p-IKKβ和Aβ1-42表达明显增多(P<0.05,P<0.01)。而空白给药组逃避潜伏期、穿越平台次数、海马Neu N表达变化差异无统计学意义,MG的i NOS表达、海马p-NF-κB p65、p-IKKβ和Aβ1-42表达明显降低,TREM2表达明显增多(P<0.05,P<0.01)。与模型组比较,YHP低剂量组逃避潜伏期明显缩短、穿越平台次数明显增加(P<0.05,P<0.01);YHP高剂量组逃避潜伏期明显缩短(P<0.05);YHP各给药组海马Neu N表达显著增加,MG的i NOS表达显著降低,Arg-1表达显著增加;血清IL-1β含量显著降低,TGF-β1含量显著增加;海马TREM2表达显著增加,p-NF-κB p65、p-IKKβ和Aβ1-42表达显著降低(P<0.01)。结论:YHP早期干预可能通过调控TREM2/NF-κB信号通路,促进海马MG由M1型向M2型转化,降低神经炎症因子释放,保护海马神经元并减少Aβ1-42沉积,最终延缓SAMP8小鼠学习记忆减退的发生。
Objective:To investigate the impact of early intervention with Yishen Huazhuo prescription(YHP)on the learning and memory of accelerated aging model mice,as well as its underlying mechanism.Method:Forty-eight 3-month-old male SAMP8 mice were randomly assigned into four groups,including the model group,low-dose YHP group,high-dose YHP group,and donepezil group.Additionally,24 SAMR1 mice of the same age were divided into a control group and a YHP treatment control group,each consisting of 12 mice.The YHP groups received YHP at doses of 6.24 g·kg^(-1) and 12.48 g·kg^(-1),while the donepezil group was treated with donepezil at a dose of 0.65 mg·kg^(-1).The model group and control groups were given physiological saline.The mice were gavaged once daily for a duration of four weeks.Spatial learning and memory abilities of mice were assessed using the Morris water maze test.Immunofluorescence staining was employed to evaluate neuronal density as well as expression levels of M1 microglial(MG)polarization marker inducible nitric oxide synthase(iNOS)and M2 MG polarization marker arginase-1(Arg-1)in the hippocampus region.Enzymelinked immunosorbent assay(ELISA)was used to measure serum levels of pro-inflammatory factor interleukin 1β(IL-1β)and anti-inflammatory factor transforming growth factor-β1(TGF-β1).Furthermore,Western blot analysis was conducted to determine expressions of amyloidβpeptide1-42(Aβ1-42)along with triggering receptor expressed on myeloid cells 2(TREM2)/nuclear factor kappa B(NF-κB)signaling pathway-related proteins TREM2,phospho(p)-NF-κB p65,and phospho-inhibitory kappa B kinaseβ(IKKβ)in the hippocampus.Result:Compared with the control group,the model group exhibited a significantly prolonged escape latency(P<0.01),a significant reduction in neuron-specific nuclear protein(NeuN)expression in the hippocampus,a significant increase in iNOS expression in MG,and a significant decrease in Arg-1 expression.The serum IL-1βcontent was significantly increased,while the TGF-β1 content was significantly decreased.Additionally,there was a significant decrease in TREM2 expression in the hippocampus and significant increases in p-NF-κB p65,p-IKKβ,and Aβ1-42 expressions(P<0.05,P<0.01).However,no significant changes were observed in escape latency,times of crossing the platform,and hippocampal NeuN expression in the YHP treatment control group.Conversely,iNOS expression in MG as well as the hippocampal p-NF-κB p65,p-IKKβ,and Aβ1-42 expressions were significantly decreased.Furthermore,TREM2 expression was significantly increased(P<0.05,P<0.01).In comparison to the model group,the low-dose YHP group showed a significantly shortened escape latency and an increased number of crossing the platform(P<0.05,P<0.01).In the high-dose YHP group,the escape latency was significantly shortened(P<0.05).In the low-dose YHP group,high-dose YHP group,the expression of NeuN in the hippocampus was significantly increased,the expression of iNOS in MG was significantly decreased,and the expression of Arg-l was significantly increased.The serum IL-1βcontent was significantly decreased,while the TGF-β1 content was significantly increased.Furthermore,the expression of TREM2 in the hippocampus was significantly increased,and the expressions of p-NF-κB p65,p-IKKβ,and Aβ1-42 were significantly decreased(P<0.01).Conclusion:Early YHP intervention may promote the transformation of hippocampal MG from M1 to M2 by regulating the TREM2/NF-κB signaling pathway,reduce the release of neuroinflammatory factors,protect hippocampal neurons,and reduce the deposition of Aβ1-42,and finally delay the occurrence of learning and memory decline in SAMP8 mice.
作者
昝树杰
王凯
徐家淳
孙伟明
倪道艳
张琳琳
刘爽
马妍
许蓬娟
李霖
ZAN Shujie;WANG Kai;XU Jiachun;SUN Weiming;NI Daoyan;ZHANG Linlin;LIU Shuang;MA Yan;XU Pengjuan;LI Lin(Tianjin University of Traditional Chinese Medicine(TCM),Tianjin 300167,China;The Second Affiliated Hospital of Tianjin University of TCM,Tianjin 300250,China)
出处
《中国实验方剂学杂志》
CAS
CSCD
北大核心
2024年第8期91-99,共9页
Chinese Journal of Experimental Traditional Medical Formulae
基金
国家中医药管理局国医大师传承工作室建设项目
国家自然科学基金项目(82104801,81973797,82274318)。
